Blood vessels and nerves develop in parallel and their survival and function in postnatal tissues are interdependent; thus, when one system is damaged, the other degenerates. Ischemic stroke is one example in which vascular damage leads to neurological degeneration and functional deficits; stroke affects 1 in 59 adults annually of whom ~5 million are permanently disabled. While the initial damage from stroke produces neuronal cell loss, the process quickly evolves into loss of other cell types and extracellular matrix, resulting in a cavitational void. Our preliminary animal studies, in a model that mimics human stroke, suggest that transplantation of neural stem cells (NSC) alone may ameliorate functional deficits caused by stroke; however, we found no neural restoration since transplanted cells integrated only into areas that retained tissue architecture. Moreover, engrafted NSC did not persist, limiting repair. We will circumvent these current limitations of cell transplantation by bioengineering a microenvironment that will sustain NSC and enable their propagation ex vivo, as well as in vivo upon transplantation. We laid the experimental groundwork for our project in previous studies in which we established a 3D model of the NSC niche via imaging and quantitative analysis, and developed biomaterials suitable for engineering this microenvironment ex vivo. We also established proof of principle that transplantation of cell-matrix constructs into stroke models is feasible and reduces lesion size. In the proposed studies, we will continue to optimize the design of our engineered niches based on our biological studies of the regulation of neurogenesis and angiogenesis in the brain (Aim 1), and by sequential testing in vitro (Aim 2) and in vivo (Aim 3) in progressively more challenging and realistic models of stroke, which will enable us to move closer to developing neurovascular regenerative therapies for human patients. Although our initial clinical target will be stroke-injured tissues, the insights gained, and strategies developed, from our proposed studies will be broadly applicable to repair of other neurovascular injuries such as traumatic brain injury and multiple sclerosis.